Computational Analysis of Cyclooxygenase Inhibition: Energetics and Dynamics

Abstract

The cyclooxygenase (COX) enzyme catalyzes the bis-dioxygenation and cyclization of arachidonic acid to form prostaglandin H2, the common precursor of bioactive prostaglandins. The genome codes two 60% sequence-identical COX isoforms. COX-1 is constitutively expressed and is associated with maintenance of gastric mucosal integrity. Selective inhibitors of COX-2, which is up-regulated at inflammation sites, are non-steroidal anti-inflammatory drugs (NSAIDs) that are less ulcerogenic than non-selective NSAIDs such as aspirin. Unfortunately, long term administration of current COX-2 selective NSAIDs carries increased risk for cardiovascular events. Amide derivatives of the non-selective inhibitor indomethacin represent a new series of COX-2 selective NSAIDs, but mutagenesis of active site residues does not reveal isoform-specific binding interactions for indomethacin amides. In Chapter II, I predict the binding mode of the enantioselective á‑substituted indomethacin ethanolamides, and I provide an energetic explanation for the increased potency of the S enantiomers against COX-1. Residue 472 is leucine in COX-2 and methionine in COX-1. The Met-472 mutation of COX-2 slows the association rate for indomethacin amides, reducing their potency. Intriguingly, in both COX isoforms residue 472 neither contacts bound ligands nor alters enzyme structure. Chapter III reveals a subtle molecular mechanism by which Met-472 restricts active site dynamic flexibility and slows the association rate of indomethacin amides. Chapter IV explores mechanisms by which COX controls the regio- and stereoselectivity of the reaction's second oxygenation, and considers routes by which prostaglandin G2 might transit to the peroxidase site for final reduction to prostaglandin H2. The putative transit channels observed in the simulations may partly explain the cooperative functioning of the COX homodimer. These studies provide additional insight into basic mechanistic details of the COX enzymes and provide rational explanations for inhibitor isoform selectivity and binding kinetic differences. This new insight may ultimately aid in design of new-generation COX inhibitors with potential therapeutic utility.